Differentiation of Murine NK Cells into Distinct Subsets Based on Variable Expression of the IL-12Rβ2 Subunit

Abstract

The cytokine IL-12 manifests its biological activity via interaction with a heterodimeric receptor (IL-12R) present on activated T and NK cells. The cDNAs for two IL-12R subunits have been cloned from human and mouse and designated IL-12Rβ1 and IL-12Rβ2. The expression of IL-12Rβ2 on T cells is influenced by cytokines, particularly IL-4, IL-12, and IFN-γ; however, little is known regarding regulation of IL-12R expression on NK cells. In this study we show that murine NK cells differentiate into IL-12Rβ2low and IL-12Rβ2high subsets after in vitro stimulation with IL-2 in the absence of exogenous polarizing cytokines. Subset development occurs gradually as NK cells expand in vitro and is generally complete by 8–12 days of culture. Once established, IL-12Rβ2low and IL-12Rβ2high subsets are highly stable in vitro and can be maintained for at least 20 days after FACS sorting. Formation of these NK subsets appears to be strain independent. Flow cytometric analyses demonstrate that both subsets express a number of NK-associated markers, including NK1.1, DX-5, Ly-49A, and Ly-49C, but that the Ly-49G2 class I inhibitory receptor is expressed predominantly on the IL-12Rβ2high population. Both IL-12Rβ2low and IL-12Rβ2high NK cells respond to exogenous IL-12 by rapid production of high levels of IFN-γ and increased lytic activity against NK-sensitive YAC-1 target cells. Analyses of cytokine gene expression by RNase protection assay indicated that similar to the recently described human NK1 subset, both IL-12Rβ2high and IL-12Rβ2low murine NK subsets expressed high levels of IFN-γ, whereas neither subset expressed mRNA for the NK2-associated cytokines IL-5 and IL-13.